Li Bing, Tian Lihua, Diao Yumei, Li Xue, Zhao Liying, Wang Xuefeng
Department of Ophthalmology, First Hospital of Liaoning Medical College, Jinzhou, Liaoning 121001, P.R. China.
Department of Otolaryngology, First Hospital of Liaoning Medical College, Jinzhou, Liaoning 121001, P.R. China.
Mol Med Rep. 2014 Jun;9(6):2245-50. doi: 10.3892/mmr.2014.2073. Epub 2014 Mar 27.
The aim of the present study was to examine the effect of exogenous IL-10 transfected rat dendritic cells (DCs) in corneal allografts. Rat lymphocyte separation medium and a cytokine induction method was used to extract and culture precursor cells of rat bone marrow-derived dendritic cells. A corneal transplant model was established, with Sprague-Dawley (SD) rats as the recipients and Wistar rats as the donors. Flow cytometry (FCM) was used to detect the expression of CD83, which indicates a mature dendritic cell, and a specific cell surface stimulatory molecule CD86. Western blot analysis was used to detect interleukin (IL)-10 protein expression and RT-PCR was used to detect cytokine IL-10, IL-2 and TGF-β mRNA expression in each group. Compared with the other groups, the survival time of corneal grafts in the IL-10-green fluorescent protein (GFP)-DC group was significantly prolonged and H&E staining demonstrated mild graft edema and inflammatory cell infiltration. There was a high expression of IL-10 and a low expression of the surface antigens, CD83 and CD86, with a lower proliferation of T lymphocytes in the IL-10-GFP-DC group. The expression of IL-10 and TGF-β in the IL-10-GFP-DC group was higher than that in the other groups, while the expression of IL-2 was lower. The transfection of the IL-10 gene inhibited dendritic cell maturation. IL-10 gene-modified immature dendritic cells (imDC) were able to inhibit corneal allograft rejection and induce immune tolerance to prolong the survival time of the corneal graft. The Th1/Th2 deviation and the high expression of TGF-β may lead to immune tolerance.
本研究的目的是检测外源性白细胞介素-10(IL-10)转染的大鼠树突状细胞(DCs)在角膜同种异体移植中的作用。采用大鼠淋巴细胞分离液和细胞因子诱导法提取并培养大鼠骨髓源性树突状细胞的前体细胞。建立角膜移植模型,以Sprague-Dawley(SD)大鼠为受体,Wistar大鼠为供体。采用流式细胞术(FCM)检测成熟树突状细胞标志物CD83及特异性细胞表面刺激分子CD86的表达。采用蛋白质印迹分析检测白细胞介素(IL)-10蛋白表达,采用逆转录-聚合酶链反应(RT-PCR)检测各组细胞因子IL-10、IL-2和转化生长因子-β(TGF-β)mRNA表达。与其他组相比,IL-10-绿色荧光蛋白(GFP)-DC组角膜移植的存活时间显著延长,苏木精-伊红(H&E)染色显示移植片轻度水肿和炎性细胞浸润。IL-10-GFP-DC组IL-10表达高,表面抗原CD83和CD86表达低,T淋巴细胞增殖较低。IL-10-GFP-DC组IL-10和TGF-β的表达高于其他组,而IL-2的表达较低。IL-10基因转染抑制树突状细胞成熟。IL-10基因修饰的未成熟树突状细胞(imDC)能够抑制角膜同种异体移植排斥反应并诱导免疫耐受,从而延长角膜移植的存活时间。Th1/Th2偏移和TGF-β的高表达可能导致免疫耐受。